The goal of this proposal is to determine how cells integrate metabolic pathways into regulatory networks. In Escherichia coli, the gcvTHP (gcv) operon and gcvB gene are part of a complex network. The gcv operon encodes the glycine cleavage (GCV) enzyme complex that provides one-carbon units for cellular methylation reactions. The gcvB gene encodes two small non-translated RNAs that in turn regulate other pathways (e.g., oppA and dppA, encoding the oligopeptide and dipeptide periplasmic binding proteins, respectively). Both gcv and gcvB are activated by the GcvA protein in response to glycine and repressed by the GcvA + GcvR proteins in the absence of glycine. Because expression of gcvB follows a pattern of regulation qualitatively similar to the gcv operon, we hypothesize that the GcvB RNAs play a role in an integrated cellular response to regulate other genes in conjunction with the GCV enzyme system. Using genetics, physiology and biochemistry, we will address two major questions. First, we will examine regulation of the gcvB gene in response to environmental signals and characterize the two RNA transcripts produced with respect to their relative amounts, stabilities and secondary structures. Second, we will determine the scope of the regulatory network and the molecular mechanisms of control by the GcvB RNAs. [unreadable] [unreadable]